The filoviruses, Ebola virus (EBOV) and Marburg virus (MARV), are important human pathogens with case fatality rates ranging from 40% to 90% for EBOV and up to 90% for MARV. Filoviruses are classified as Category A Priority Pathogens by the NIAID/NIH and CDC, and there are presently no approved active or passive interventions for exposure resulting from natural outbreak, laboratory accident, or deliberate misuse. Public health concern is based on both the emerging infectious disease status of these viruses and their potential use as biologic weapons. An effective post-exposure vaccine would find application with medical personnel and close contacts during outbreaks in endemic areas of Africa, with laboratory workers engaged in filovirus research, and with military and civilian personnel threatened by weaponized filoviruses. The ideal vaccine to meet both the outbreak and bio-weapon scenarios would confer post-exposure protection against all species of EBOV and MARV with a single administration. Among the vaccine technologies investigated to date, a tri-valent filovirus vaccine vectored by recombinant vesicular stomatitis virus (rVSV) has shown the greatest potential as a single administration vaccine with the capacity to rapidly provide broad post-exposure protection against EBOV and MARV. Profectus BioSciences has developed a replication competent attenuated rVSV vaccine delivery platform (rVSVN4CT1) that: 1) retains the immunogenicity of vaccines based on the non-attenuated vector, 2) has been manufactured under cGMPs at commercial scale, and 3) has been shown to be safe and immunogenic in multiple clinical trials. Thus, this application proposes: 1) examine 4 rVSV vector designs to determine which provides the best combination of protective efficacy, stability, and manufacturing yield, 2) to confirm that an attenuated trivalent rVSVN4CT1 vectored filovirus vaccine will provide post-exposure protection of monkeys against challenge with filoviruses, 3) compliantly prepare seed stocks of the three viruses in the vaccine under conditions that will support future manufacture under cGMPs, 4) use the compliant seed stocks to prepare a tri-valent vaccine that will be tested in a GLP neuro-toxicology study in nonhuman primates (NHPs) to confirm safety, 5) use the compliant tri-valent vaccine in development studies to identify a lyophilized formulation with the stability characteristics required for practical field use, 6) test the lyophilized tri-valent vaccine to confirm that it protects NHPs from filoviruses when administered and postexposure, and 7) determine if tri-valent vaccine will synergize with small interfering RNA (siRNA) and/or monoclonal antibodies in postexposure protection from RPI and RP2.